Kinase inhibitor and method for treatment of related diseases

ABSTRACT

Disclosed is a compound of (aminophenylamino) pyrimidyl benzamides and a synthesis method thereof. The compound has Btk-inhibition activity and can be used to treat autoimmune diseases, heteroimmune diseases, cancers or thromboembolic diseases.

FIELD OF THE INVENTION

The present application provides the molecular structures of compoundsof (aminophenylamino) pyrimidyl benzamides and synthesis methodsthereof, as well as use of the compounds in inhibiting kinases andtreating B-cell associated diseases.

BACKGROUND OF THE INVENTION

The kinase's action mechanism is to transfer phosphate groups fromhigh-energy donor molecules (e.g., ATP) to specific molecules, which isa process called phosphorylation. Protein kinases alter the activitiesof specific proteins through phosphorylation so as to control andregulate protein-associated signal transduction and other effects oncells. Due to the importance of protein kinases in cell signaling, theselectivity of some small molecule compounds for specific kinases willbe helpful for further understanding on the cell signaling process.Meanwhile, small molecule compounds control the functions of cells bymodulating the activities of kinases, which makes protein kinases becomegood drug targets in the treatment of clinical diseases.

Bruton's tyrosine kinase (Btk), a member of the Tec family ofnon-receptor tyrosine kinases, plays a key role in signal transductionin hematopoietic cells (except T lymphocytes and plasma cells),especially in the B cells which play an important role in thepathogenesis of autoimmune and inflammatory diseases. Btk has shown goodclinical efficacy in many serious refractory diseases, such asrheumatoid arthritis, lymphoma and leukemia.

Btk plays a critical role in the process of B-cell development,differentiation, proliferation, activation and survival. The effect ofBtk on B cells is achieved by controlling B-cell receptor (BCR)signaling pathway. Btk locates at adjacent downstream of the BCR. Btkpasses down the signal upon BCR stimulation, and after a series ofsignal transduction, finally leading to intracellular calciummobilization and protein kinase C activation. X-linkedagammaglobulinemia (also called Bruton's syndrome, XLA) is a raregenetic disease. These XLA patients are unable to produce mature Bcells. Normal B cells resist external infection by producing antibodies(called immunoglobulins). Due to the lack of B cells and antibodies, XLApatients are easy to obtain serious or even fatal infections. Furtherresearches found that the direct reason that inhibits B-cell developmentis gene mutation of Btk. Thus it is proved that Btk plays an extremelyimportant role in the development and function of normal B cells.

Btk becomes a remarkable drug target in cancers that are relevant to theB-cell, especially the B-cell lymphoma and leukemia.

Cells need BCR signals to grow and proliferate. Since Btk is anindispensable key member in the BCR signaling pathway, Btk inhibitorscan block BCR signaling and induce apoptosis of cancer cells. Currently,there are two Btk inhibitors in the United States and Europe forclinical treatment of chronic lymphocytic leukemia (Cll) and smalllymphocytic lymphoma (Sll): PCI-32765 (clinical phase III) and AVL-292(clinical phase I). (See S E Herman et al. (2011), Blood 117 (23):6287-96). Btk is also associated with acute lymphoblastic leukemia.Acute lymphoblastic leukemia is the most common cancer in children, andhas a poor prognosis in adult patients. Genetic analysis found that thedeficiency of BTK expression was found in all types of leukemia.Defective Btk protects leukemia cells from apoptosis.

Btk is also a therapeutic target for autoimmune diseases. Rheumatoidarthritis is a chronic autoimmune disease. Btk is an important componentof BCR signaling in B cells and FC-γ signaling in bone marrow cells. Btkinhibitors are expected to reduce two main components of autoimmunediseases: pathogenic auto-antibodies produced by B cells andpro-inflammatory cytokine produced by myeloid cells. In cellexperiments, it is proved that Btk inhibitors can effectively reduceauto-antibodies and pro-inflammatory cytokines. In mice withcollagen-induced arthritis, Btk inhibitors reduced in vivo level ofauto-antibodies and effectively controlled the disease. These resultsprovide a new understanding of Btk functions during the development ofB-cells or bone-marrow-cells driven diseases, and provide a convincingreason for targeting Btk in the treatment of rheumatoid arthritis. (SeeL A Honigberg et al. (2010), Proc Natl Acad Sci USA 107 (29): 13075-80.J A Di Paolo et al. (2011), Nat Chem Biol 7 (1): 41-50.)

The role of Btk in inflammatory diseases has been demonstrated by a ratbasophilic leukemia cells (RBL-2H3) model. RBL-2H3 is a common model formast cell inflammatory diseases research. Mast cells are rich ofbasophilic granules, and play a leading role in immunoglobulin E(IgE)-mediated allergic reactions. Small interfering RNA (siRNA), andLFM-A13 (an effective Btk inhibitor) can suppress the mast cell inducedinflammatory response by inhibiting Btk activity. In the mast cellstreated with siRNA and LFM-A13, the release of a pro-inflammatorymediator, histamine, is reduced by 20-25%.

It is also reported in literatures that Btk is used as a therapeutictarget in heteroimmune diseases and thromboembolic diseases.

Therefore, the present disclosure aims to provide a novel compound fortreating autoimmune diseases, heteroimmune diseases, inflammatorydiseases, cancers, or thromboembolic diseases.

SUMMARY OF THE INVENTION

In one aspect of the present disclosure, it provides:

A compound of formula (I),

or pharmaceutically acceptable salts thereof, wherein:

W is selected from H, C₁₋₆ alkyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃,and —(NH—CO)_(n)—NH-L-L₃;

wherein:

L is a bond, C₁₋₃ alkylene or C₂₋₃ alkenylene;

L₃ is C₃₋₈ cycloalkyl, such as

Aryl such as phenyl, naphthyl, phenanthryl, anthryl, fluorenyl, andindenyl, or heteroaryl such as

The C₃₋₈ cycloalkyl, aryl and heteroaryl is optionally substituted with1, 2 or 3 substituents selected from the group consisting of halogensuch as F and Cl, amino, C₁₋₆ alkyl, C₁₋₆ alkoxyl, halo-C₁₋₆ alkyl suchas perhalo-C₁₋₆ alkyl such as CF₃;

n is an integer of 0 or 1;

X is selected from H, halogen such as F and Cl, and C₁₋₆ alkyl such asmethyl;

R₁ and R₂, same or different from each other, are each independentlyselected from H, C(O) and S(O)₂;

L₁ and L₂, same or different from each other, are each independentlyselected from C₂₋₃ alkenyl optionally substituted with C₁₋₃ alkyl, andC₁₋₃ alkyl-NHC(O)—C₂₋₃ alkenyl;

with the provisos that when R₁ is H, L₁ is not present; and when R₂ isH, L₂ is not present.

In a preferred embodiment,

W is selected from H, ethyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and—(NH—CO)_(n)—NH-L-L₃,

wherein:

L is a bond or vinylene;

L₃ is cyclopropyl, phenyl, naphthyl, isoxazolyl or benzo[d][1,3]dioxolegroup optionally substituted with 1 or 2 substituents selected from F,Cl, amino, methoxyl and CF₃;

n is an integer of 1.

In another preferred embodiment,

X is selected from H, F, Cl, and methyl.

In another preferred embodiment,

R₁ and R₂, same or different from each other, are each independentlyselected from H, C(O) and S(O)₂;

L₁ and L₂, same or different from each other, are each independentlyselected from C₂₋₃ alkenyl, and methyl-NHC(O)-ethenyl;

with the provisos that when R₁ is H, L₁ is not present; and when R₂ isH, L₂ is not present.

In another preferred embodiment,

W is selected from H, ethyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and—(NH—CO)_(n)—NH-L-L₃, wherein:

L is a bond or vinylene;

L₃ is cyclopropyl, phenyl, naphthyl, isoxazolyl or benzol[d][1,3]dioxolegroup optionally substituted with 1 or 2 substituents selected from F,Cl, amino, methoxyl and CF₃;

n is an integer of 1;

X is selected from H, F, Cl, and methyl;

R₁ and R₂, same or different from each other, are each independentlyselected from H, C(O) and S(O)₂;

L₁ and L₂, same or different from each other, are each independentlyselected from C₂₋₃ alkenyl, and methyl —NHC(O)-ethenyl;

with the provisos that when R₁ is H, L₁ is not present; and when R₂ isH, L₂ is not present. In another aspect of the present disclosure, itprovides a compound selected from:

In another aspect of the present disclosure, it provides pharmaceuticalcompositions comprising a therapeutically effective amount of thecompound of the present invention and pharmaceutically acceptableexcipients.

In another aspect of the present disclosure, it provides uses of thecompounds or the compositions of the present invention in themanufacture of medicaments for treating the following diseases orconditions: autoimmune diseases, heteroimmune diseases, inflammatorydiseases, cancers or thromboembolic diseases.

In another aspect of the present disclosure, it provides the compoundsor the compositions of the present invention used in methods fortreating the diseases or conditions as follows: autoimmune diseases,heteroimmune diseases, inflammatory diseases, cancers or thromboembolicdiseases.

In another aspect of the present disclosure, it provides methods fortreating diseases or conditions as follows: autoimmune diseases,heteroimmune diseases, inflammatory diseases, cancers or thromboembolicdiseases, said methods comprising administering the compounds or thecompositions of the present invention to subjects in need thereof, e.g.a mammal such as human.

For any and all of the embodiments, substituents can be selected from asubset of the listed alternatives. For example, in some embodiments, Wis selected from H, ethyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and—(NH—CO)_(n)—NH-L-L₃. In some further embodiments, W is selected from—(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and —(NH—CO)_(n)—NH-L-L₃. In somefurther embodiments, W is selected from —(NH—CO)_(n)-L-L₃.

Other objects, features and advantages of the methods and compositionsdescribed herein will become apparent from the following detaileddescription. It should be understood, however, that the detaileddescription and the specific examples, while indicating specificembodiments, are given by way of illustration only, since variouschanges and modifications within the spirit and scope of the presentdisclosure will become apparent to those skilled in the art from thisdetailed description. The section headings used herein are fororganizational purposes only and are not to be construed as limiting thesubject matter described. All documents, or portions of documents, citedin the application including, but not limited to, patents, patentapplications, articles, books, manuals, and treatises are herebyexpressly incorporated by reference in their entirety for any purpose.

EMBODIMENTS

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as they are commonly understood by one skilled inthe art to which the claimed subject matter belongs.

Definition of standard chemistry terms may be found in reference works,including Carey and Sundberg “ADVANCED ORGANIC CHEMISTRY 4^(TH) ED.”Vols. A (2000) and B (2001), Plenum Press, New York.

“C₁₋₆ alkyl” refers to an alkyl group with 1 to 6 carbon atoms,including methyl, ethyl, propyl, butyl, pentyl and hexyl, and all thepossible isomeric forms thereof, e.g., n-propyl and isopropyl, n-butyl,isobutyl, sec-butyl and tert-butyl, and the like. “C₁₋₆ alkyl” includesall sub-ranges contained therein, e.g. C₁₋₂ alkyl, C₁₋₃ alkyl, C₁₋₄alkyl, C₁₋₅ alkyl, C₂₋₅ alkyl, C₃₋₅ alkyl, C₄₋₅ alkyl, C₃₋₄ alkyl, C₃₋₅alkyl and C₄₋₅ alkyl.

“C₁₋₃ alkylene” includes methylene, ethylidene, propylidene andisopropylidene.

“C₂₋₃ alkenyl” includes ethenyl (—CH═CH₂), propenyl (—CH═CHCH₃) andisopropenyl (—C(CH₃)═CH₂).

“C₂₋₃ alkenylene” includes ethenylene (—CH═CH—), propenylene(—CH═CHCH₂—) and isopropenylene (—C(CH₃)═CH—).

The term “aromatic group” refers to a planar ring having a delocalizedmembered π-electron system containing 4n+2π electrons, where n is aninteger. Aromatic groups can be formed from five, six, seven, eight,nine or more than nine atoms. Aromatic groups can be optionallysubstituted. Aromatic groups include “aryl” (each of the atoms formingthe ring is a carbon atom), and “heteroaryl” (the atoms forming the ringinclude carbon atom(s) and heteroatom(s) selected from such as oxygen,sulfur and nitrogen). “Aryl” and “heteroaryl” include monocyclic orfused-ring polycyclic (i.e., rings which share adjacent pairs of ringatoms) groups.

Examples of aryl groups include, but are not limited to phenyl,naphthalenyl, phenanthrenyl, anthracenyl, fluorenyl and indenyl.

Examples of heteroaryl groups include,

“C₃₋₈ cycloalkyl” refers to a non-aromatic monocyclic or polycyclicradical that contains only carbon and hydrogen, having 3 to 8 carbonsforming a ring, and may be saturated, partially unsaturated, or fullyunsaturated. Examples of C₃₋₈ cycloalkyl groups include the following:

“Halogen” refers to fluoro, chloro, bromo and iodo.

“C₁₋₆ alkoxyl” refers to the group (C₁₋₆ alkyl)O—, wherein the C₁₋₆alkyl is as defined herein.

“Halo-C₁₋₆ alkyl” refers to halo-(C₁₋₆ alkyl)-, wherein the C₁₋₆ alkylis as defined herein. Halo-C₁₋₆ alkyl includes perhalogenated C₁₋₆alkyl, wherein all the hydrogen atoms in C₁₋₆ alkyl are replaced withhalogen, such as —CF₃, —CH₂CF₃, —CF₂CF₃, —CH₂CH₂CF₃ and the like.

“C₂₋₃ alkenyl optionally substituted with C₁₋₃ alkyl” refers to a C₂₋₃alkenyl or a C₂₋₃ alkenyl substituted with C₁₋₃ alkyl, wherein itconnects to the main structure of the compound through C₂₋₃ alkenyl.

“C₁₋₃ alkyl-NHC(O)—C₂₋₃ alkenyl” refers to C₂₋₃ alkenyl substituted withC₁₋₃ alkyl-NHC(O), wherein it connects to the main structure of thecompound through C₂₋₃ alkenyl.

The term “bond” refers to a chemical bond between two atoms, or twomoieties when the atoms joined by the bond are considered to be part oflarger substructure.

The term “pharmaceutically acceptable”, with respect to a formulation,composition or ingredient, as used herein, means having no persistentdetrimental effect on the general health of the subject being treated ordoes not abrogate the biological activity or properties of the compound,and is relatively nontoxic.

The term “Bruton's tyrosine kinase,” as used herein, refers to Bruton'styrosine kinase from Homo sapiens, as disclosed in, e.g., U.S. Pat. No.6,326,469 (GenBank Accession No. NP 000052).

The terms “effective amount” or “therapeutically effective amount,” asused herein, refer to a sufficient amount of an agent or a compoundbeing administered which will relieve to some extent one or more of thesymptoms of the disease or condition being treated. The result can bereduction and/or alleviation of the signs, symptoms, or causes of adisease, or any other desired alteration of a biological system. Forexample, an “effective amount” for therapeutic uses is the amount of thecomposition including a compound as disclosed herein required to providea clinically significant decrease in disease symptoms without undueadverse side effects. An appropriate “effective amount” in anyindividual case may be determined using techniques, such as a doseescalation study. The term “therapeutically effective amount” includes,for example, a prophylactically effective amount. An “effective amount”of a compound disclosed herein is an amount effective to achieve adesired pharmacologic effect or therapeutic improvement without undueadverse side effects. It is understood that “an effect amount” or “atherapeutically effective amount” can vary from subject to subject, dueto variation in metabolism of the compound, age, weight, generalcondition of the subject, the condition being treated, the severity ofthe condition being treated, and the judgment of the prescribingphysician. By way of example only, therapeutically effective amounts maybe determined by routine experimentation, including but not limited to adose escalation clinical trial.

The terms “inhibits”, “inhibiting” or “inhibitor” of a kinase, as usedherein, refer to inhibition of enzymatic phosphotransferase activity.

Autoimmune diseases, as disclosed herein, include but are not limitedto, rheumatoid arthritis, psoriatic arthritis, osteoarthritis, Still'sdisease, juvenile arthritis, lupus, diabetes, myasthenia gravis,Hashimoto's thyroiditis, Ord's thyroiditis, Graves' disease Sjögren'ssyndrome, multiple sclerosis, Guillain-Barré syndrome, acutedisseminated encephalomyelitis, Addison's disease, opsoclonus-myoclonussyndrome, ankylosing spondylitisis, antiphospholipid antibody syndrome,aplastic anemia, autoimmune hepatitis, coeliac disease, Goodpasture'ssyndrome, idiopathic thrombocytopenic purpura, optic neuritis,scleroderma, primary biliary cirrhosis, Reiter's syndrome, Takayasu'sarteritis, temporal arteritis, warm autoimmune hemolytic anemia,Wegener's granulomatosis, psoriasis, alopecia universalis, Behçet'sdisease, chronic fatigue, dysautonomia, endometriosis, interstitialcystitis, neuromyotonia, scleroderma, and vulvodynia.

Heteroimmune diseases, as disclosed herein, include but are not limitedto graft versus host disease, transplantation, transfusion, anaphylaxis,allergies (e.g., allergies to plant pollens, latex, drugs, foods, insectpoisons, animal hair, animal dander, dust mites, or cockroach calyx),type I hypersensitivity, allergic conjunctivitis, allergic rhinitis, andatopic dermatitis.

Inflammatory diseases, as disclosed herein, include but are not limitedto asthma, inflammatory bowel disease, appendicitis, blepharitis,bronchiolitis, bronchitis, bursitis, cervicitis, cholangitis,cholecystitis, colitis, conjunctivitis, cystitis, dacryoadenitis,dermatitis, dermatomyositis, encephalitis, endocarditis, endometritis,enteritis, enterocolitis, epicondylitis, epididymitis, fasciitis,fibrositis, gastritis, gastroenteritis, hepatitis, hidradenitissuppurativa, laryngitis, mastitis, meningitis, myelitis myocarditis,myositis, nephritis, oophoritis, orchitis, osteitis, otitis,pancreatitis, parotitis, pericarditis, peritonitis, pharyngitis,pleuritis, phlebitis, pneumonitis, pneumonia, proctitis, prostatitis,pyelonephritis, rhinitis, salpingitis, sinusitis, stomatitis, synovitis,tendonitis, tonsillitis, uveitis, vaginitis, vasculitis, and vulvitis.

Cancers, as disclosed herein, e.g., B-cell proliferative disorders,which include, but are not limited to diffuse large B cell lymphoma,follicular lymphoma, chronic lymphocytic lymphoma, chronic lymphocyticleukemia, B-cell prolymphocytic leukemia, lymphoplasmacyticlymphoma/Waldenström macroglobulinemia, splenic marginal zone lymphoma,plasma cell myeloma, plasmacytoma, extranodal marginal zone B celllymphoma, nodal marginal zone B cell lymphoma, mantle cell lymphoma,mediastinal (thymic) large B cell lymphoma, intravascular large B celllymphoma, primary effusion lymphoma, burkitt lymphoma/leukemia, andlymphomatoid granulomatosis.

Thromboembolic disorders, as disclosed herein, which include, but arenot limited to myocardial infarct, angina pectoris (including unstableangina), reocclusions or restenoses after angioplasty or aortocoronarybypass, stroke, transitory ischemia, peripheral arterial occlusivedisorders, pulmonary embolisms, and deep venous thromboses.

Symptoms, diagnostic tests, and prognostic tests for each of theabove-mentioned conditions are known in the art. See, e.g., Harrison'sPrinciples of Internal Medicine©,” 16^(th) ed., 2004, The McGraw-HillCompanies, Inc. Dey et al. (2006), Cytojournal 3(24), and the “RevisedEuropean American Lymphoma” (REAL) classification system (see, e.g., thewebsite maintained by the National Cancer Institute).

A number of animal models of are useful for establishing a range oftherapeutically effective doses of irreversible Btk inhibitor compoundsfor treating any of the foregoing diseases.

For example, dosing of irreversible Btk inhibitor compounds for treatingan autoimmune disease can be assessed in a mouse model of rheumatoidarthritis. In this model, arthritis is induced in Balb/c mice byadministering anti-collagen antibodies and lipopolysaccharide. SeeNandakumar et al. (2003), Am. J. Pathol 163:1827-1837.

In another example, dosing of irreversible Btk inhibitors for thetreatment of B-cell proliferative disorders can be examined in, e.g., ahuman-to-mouse xenograft model in which human B-cell lymphoma cells(e.g. Ramos cells) are implanted into immunodefficient mice (e.g.,“nude” mice) as described in, e.g., Pagel et al. (2005), Clin Cancer Res11(13):4857-4866.

Animal models for treatment of thromboembolic disorders are also known.

The therapeutic efficacy of the compound for one of the foregoingdiseases can be optimized during a course of treatment. For example, asubject being treated can undergo a diagnostic evaluation to correlatethe relief of disease symptoms or pathologies to inhibition of in vivoBtk activity achieved by administering a given dose of an irreversibleBtk inhibitor. Cellular assays known in the art can be used to determinein vivo activity of Btk in the presence or absence of an irreversibleBtk inhibitor. For example, since activated Btk is phosphorylated attyrosine 223 (Y223) and tyrosine 551 (Y551), phospho-specificimmunocytochemical staining of P-Y223 or P-Y551-positive cells can beused to detect or quantify activation of Bkt in a population of cells(e.g., by FACS analysis of stained vs unstained cells). See, e.g.,Nisitani et al. (1999), Proc. Natl. Acad. Sci, USA 96:2221-2226. Thus,the amount of the Btk inhibitor compound that is administered to asubject can be increased or decreased as needed so as to maintain alevel of Btk inhibition optimal for treating the subject's diseasestate.

The starting material used for the synthesis of the compounds describedherein may be synthesized or can be obtained from commercial sources,such as, but not limited to, Aldrich Chemical Co. (Milwaukee, Wis.),Bachem (Torrance, Calif.), or Sigma Chemical Co. (St. Louis, Mo.). Thecompounds described herein, and other related compounds having differentsubstituents can be synthesized using techniques and materials known tothose of skill in the art, such as described, for example, in March,ADVANCED ORGANIC CHEMISTRY 4^(th) Ed., (Wiley 1992); Carey and Sundberg,ADVANCED ORGANIC CHEMISTRY 4^(th) Ed., Vols. A and B (Plenum 2000,2001); Green and Wuts, PROTECTIVE GROUPS IN ORGANIC SYNTHESIS 3^(rd)Ed., (Wiley 1999); Fieser and Fieser's Reagents for Organic Synthesis,Volumes 1-17 (John Wiley and Sons, 1991); Rodd's Chemistry of CarbonCompounds, Volumes 1-5 and Supplementals (Elsevier Science Publishers,1989); Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991); andLarock's Comprehensive Organic Transformations (VCH Publishers Inc.,1989). (all of which are incorporated by reference in their entirety).Other methods for the synthesis of compounds described herein may befound in International Patent Publication No. WO 01/01982901, Arnold etal. Bioorganic & Medicinal Chemistry Letters 10 (2000) 2167-2170;Burchat et al. Bioorganic & Medicinal Chemistry Letters 12 (2002)1687-1690. General methods for the preparation of compound as disclosedherein may be derived from known reactions in the field, and thereactions may be modified by the use of appropriate reagents andconditions, as would be recognized by the skilled person, for theintroduction of the various moieties found in the formulae as providedherein. As a guide the following synthetic methods may be utilized.

The products of the reactions may be isolated and purified, if desired,using conventional techniques, including, but not limited to,filtration, distillation, crystallization, chromatography and the like.Such materials may be characterized using conventional means, includingphysical constants and spectral data.

Compounds described herein may be prepared using the synthetic methodsdescribed herein as a single isomer or a mixture of isomers.

The compounds described herein may possess one or more stereocenters andeach center may exist in the R or S configuration. The compoundspresented herein include all diastereomeric, enantiomeric, and epimericforms as well as the appropriate mixtures thereof. Stereoisomers may beobtained, if desired, by methods known in the art as, for example, theseparation of stereoisomers by chiral chromatographic columns.

Diasteromeric mixtures can be separated into their individualdiastereomers on the basis of their physical chemical differences bymethods known, for example, by chromatography and/or fractionalcrystallization. In one embodiment, enantiomers can be separated bychiral chromatographic columns. In other embodiments, enantiomers can beseparated by converting the enantiomeric mixture into a diastereomericmixture by reaction with an appropriate optically active compound (e.g.,alcohol), separating the diastereomers and converting (e.g.,hydrolyzing) the individual diastereomers to the corresponding pureenantiomers. All such isomers, including diastereomers, enantiomers, andmixtures thereof are considered as part of the compositions describedherein.

The methods and formulations described herein include the use ofN-oxides, crystalline forms (also known as polymorphs), orpharmaceutically acceptable salts of compounds described herein, as wellas active metabolites of these compounds having the same type ofactivity. In some situations, compounds may exist as tautomers. Alltautomers are included within the scope of the compounds presentedherein. In addition, the compounds described herein can exist inunsolvated as well as solvated forms with pharmaceutically acceptablesolvents such as water, ethanol, and the like. The solvated forms of thecompounds presented herein are also considered to be disclosed herein.

Compounds in unoxidized form can be prepared from N-oxides by treatingwith a reducing agent, such as, but not limited to, sulfur, sulfurdioxide, triphenyl phosphine, lithium borohydride, sodium borohydride,phosphorus trichloride, tribromide, or the like in a suitable inertorganic solvent, such as, but not limited to, acetonitrile, ethanol,aqueous dioxane, or the like at 0 to 80° C.

In some embodiments, compounds described herein are prepared asprodrugs. A “prodrug” refers to an agent that is converted into theparent drug in vivo. Prodrugs are often useful because, in somesituations, they may be easier to administer than the parent drug. Theymay, for instance, be bioavailable by oral administration whereas theparent is not. The prodrug may also have improved solubility inpharmaceutical compositions over the parent drug. An example, withoutlimitation, of a prodrug would be a compound described herein, which isadministered as an ester (the “prodrug”) to facilitate transmittalacross a cell membrane where water solubility is detrimental to mobilitybut which then is metabolically hydrolyzed to the carboxylic acid, theactive entity, once inside the cell where water-solubility isbeneficial. A further example of a prodrug might be a short peptide(polyaminoacid) bonded to an acid group where the peptide is metabolizedto reveal the active moiety. In certain embodiments, upon in vivoadministration, a prodrug is chemically converted to the biologically,pharmaceutically or therapeutically active form of the compound. Incertain embodiments, a prodrug is enzymatically metabolized by one ormore steps or processes to the biologically, pharmaceutically ortherapeutically active form of the compound. To produce a prodrug, apharmaceutically active compound is modified such that the activecompound will be regenerated upon in vivo administration. The prodrugcan be designed to alter the metabolic stability or the transportcharacteristics of a drug, to mask side effects or toxicity, to improvethe flavor of a drug or to alter other characteristics or properties ofa drug. By virtue of knowledge of pharmacodynamic processes and drugmetabolism in vivo, those of skill in this art, once a pharmaceuticallyactive compound is known, can design prodrugs of the compound. (see, forexample, Nogrady (1985) Medicinal Chemistry A Biochemical Approach,Oxford University Press, New York, pages 388-392; Silverman (1992), TheOrganic Chemistry of Drug Design and Drug Action, Academic Press, Inc.,San Diego, pages 352-401, Saulnier et al., (1994), Bioorganic andMedicinal Chemistry Letters, Vol. 4, p. 1985).

Prodrug forms of the herein described compounds, wherein the prodrug ismetabolized in vivo to produce a derivative as set forth herein areincluded within the scope of the claims. In some cases, some of theherein-described compounds may be a prodrug for another derivative oractive compound.

Prodrugs are often useful because, in some situations, they may beeasier to administer than the parent drug. They may, for instance, bebioavailable by oral administration whereas the parent is not. Theprodrug may also have improved solubility in pharmaceutical compositionsover the parent drug. Prodrugs may be designed as reversible drugderivatives, for use as modifiers to enhance drug transport tosite-specific tissues. In some embodiments, the design of a prodrugincreases the effective water solubility. See, e.g., Fedorak et al., Am.J. Physiol, 269:G210-218 (1995); McLoed et al., Gastroenterol,106:405-413 (1994); Hochhaus et al., Biomed. Chrom., 6:283-286 (1992);J. Larsen and H. Bundgaard, Int. J. Pharmaceutics, 37, 87 (1987); J.Larsen et al., Int. J. Pharmaceutics, 47, 103 (1988); Sinkula et al., J.Pharm. Sci., 64:181-210 (1975); T. Higuchi and V. Stella, Pro-drugs asNovel Delivery Systems, Vol. 14 of the A.C.S. Symposium Series; andEdward B. Roche, Bioreversible Carriers in Drug Design, AmericanPharmaceutical Association and Pergamon Press, 1987, all incorporatedherein in their entirety.

Compounds described herein include isotopically-labeled compounds, whichare identical to those recited in the various formulas and structurespresented herein, but for the fact that one or more atoms are replacedby an atom having an atomic mass or mass number different from theatomic mass or mass number usually found in nature. Examples of isotopesthat can be incorporated into the present compounds include isotopes ofhydrogen, carbon, nitrogen, oxygen, fluorine and chlorine, such as ²H,³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, ¹⁷O, ³⁵S, ¹⁸F, ³⁶Cl, respectively. Certainisotopically-labeled compounds described herein, for example those intowhich radioactive isotopes such as ³H and ¹⁴C are incorporated, areuseful in drug and/or substrate tissue distribution assays. Further,substitution with isotopes such as deuterium, i.e., ²H, can affordcertain therapeutic advantages resulting from greater metabolicstability, for example increased in vivo half-life or reduced dosagerequirements.

In additional or further embodiments, the compounds described herein aremetabolized upon administration to an organism in need to produce ametabolite that is then used to produce a desired effect, including adesired therapeutic effect.

Compounds described herein may be formed as, and/or used as,pharmaceutically acceptable salts. The type of pharmaceutical acceptablesalts, include, but are not limited to: (1) acid addition salts, formedby reacting the free base form of the compound with a pharmaceuticallyacceptable inorganic acid such as hydrochloric acid, hydrobromic acid,sulfuric acid, nitric acid, phosphoric acid, metaphosphoric acid, andthe like; or with an organic acid such as acetic acid, propionic acid,hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid,lactic acid, malonic acid, succinic acid, malic acid, maleic acid,fumaric acid, trifluoroacetic acid, tartaric acid, citric acid, benzoicacid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid,2-hydroxyethanesulfonic acid, benzenesulfonic acid, toluenesulfonicacid, 2-naphthalenesulfonic acid,4-methylbicyclo-[2.2.2]oct-2-ene-1-carboxylic acid, glucoheptonic acid,4,4′-methylenebis-(3-hydroxy-2-ene-1-carboxylic acid), 3-phenylpropionicacid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfuricacid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylicacid, stearic acid, muconic acid, and the like; (2) salts formed when anacidic proton present in the parent compound either is replaced by ametal ion, e.g., an alkali metal ion (e.g. lithium, sodium, potassium),an alkaline earth ion (e.g. magnesium, or calcium), or an aluminum ion;or coordinates with an organic base. Acceptable organic bases includeethanolamine, diethanolamine, triethanolamine, tromethamine,N-methylglucamine, and the like. Acceptable inorganic bases includealuminum hydroxide, calcium hydroxide, potassium hydroxide, sodiumcarbonate, sodium hydroxide, and the like.

The corresponding counterions of the pharmaceutically acceptable saltsmay be analyzed and identified using various methods including, but notlimited to, ion exchange chromatography, ion chromatography, capillaryelectrophoresis, inductively coupled plasma, atomic absorptionspectroscopy, mass spectrometry, or any combination thereof.

The salts are recovered by using at least one of the followingtechniques: filtration, precipitation with a non-solvent followed byfiltration, evaporation of the solvent, or, in the case of aqueoussolutions, lyophilization.

It should be understood that a reference to a pharmaceuticallyacceptable salt includes the solvent addition forms or crystal formsthereof, particularly solvates or polymorphs. Solvates contain eitherstoichiometric or non-stoichiometric amounts of a solvent, and may beformed during the process of crystallization with pharmaceuticallyacceptable solvents such as water, ethanol, and the like. Hydrates areformed when the solvent is water, or alcoholates are formed when thesolvent is alcohol. Solvates of compounds described herein can beconveniently prepared or formed during the processes described herein.In addition, the compounds provided herein can exist in unsolvated aswell as solvated forms. In general, the solvated forms are consideredequivalent to the unsolvated forms for the purposes of the compounds andmethods provided herein.

It should be understood that a reference to a salt includes the solventaddition forms or crystal forms thereof, particularly solvates orpolymorphs. Solvates contain either stoichiometric or non-stoichiometricamounts of a solvent, and are often formed during the process ofcrystallization with pharmaceutically acceptable solvents such as water,ethanol, and the like. Hydrates are formed when the solvent is water, oralcoholates are formed when the solvent is alcohol. Polymorphs includethe different crystal packing arrangements of the same elementalcomposition of a compound. Polymorphs usually have different X-raydiffraction patterns, infrared spectra, melting points, density,hardness, crystal shape, optical and electrical properties, stability,and solubility. Various factors such as the recrystallization solvent,rate of crystallization, and storage temperature may cause a singlecrystal form to dominate.

Compounds described herein may be in various forms, including but notlimited to, amorphous forms, milled forms and nano-particulate forms. Inaddition, compounds described herein include crystalline forms, alsoknown as polymorphs. Polymorphs include the different crystal packingarrangements of the same elemental composition of a compound. Polymorphsusually have different X-ray diffraction patterns, infrared spectra,melting points, density, hardness, crystal shape, optical and electricalproperties, stability, and solubility. Various factors such as therecrystallization solvent, rate of crystallization, and storagetemperature may cause a single crystal form to dominate.

The screening and characterization of the pharmaceutically acceptablesalts, polymorphs and/or solvates may be accomplished using a variety oftechniques including, but not limited to, thermal analysis, x-raydiffraction, spectroscopy, vapor sorption, and microscopy. Thermalanalysis methods address thermo chemical degradation or thermo physicalprocesses including, but not limited to, polymorphic transitions, andsuch methods are used to analyze the relationships between polymorphicforms, determine weight loss, to find the glass transition temperature,or for excipient compatibility studies. Such methods include, but arenot limited to, Differential scanning calorimetry (DSC), ModulatedDifferential Scanning calorimetry (MDCS), Thermogravimetric analysis(TGA), and Thermogravimetric and Infrared analysis (TG/IR). X-raydiffraction methods include, but are not limited to, single crystal andpowder diffractometers and synchrotron sources. The variousspectroscopic techniques used include, but are not limited to, Raman,FTIR, UVIS, and NMR (liquid and solid state). The various microscopytechniques include, but are not limited to, polarized light microscopy,Scanning Electron Microscopy (SEM) with Energy Dispersive X-Ray Analysis(EDX), Environmental Scanning Electron Microscopy with EDX (in gas orwater vapor atmosphere), IR microscopy, and Raman microscopy.

Throughout the specification, groups and substituents thereof can bechosen by one skilled in the field to provide stable moieties andcompounds.

EXAMPLES

The following specific and non-limiting examples are to be construed asmerely illustrative, and do not limit the present disclosure in any waywhatsoever. Without further elaboration, it is believed that one skilledin the art can, based on the description herein, utilize the presentdisclosure to its fullest extent.

Synthesis of Compounds

Synthetic Scheme I

Step 1:

m-phenylenediamine (0.500 g, 4.62 mmol), (Boc)₂O (0.92 mL, 4.02 mmol)and triethylamine (1.4 mL, 9.98 mmol) were added to a mixed solventsystem of 1,4-dioxane and water (30 mL, 2:1 V/V) that has been cooled to0° C. After stirring for 1 hour at 0° C., the reaction system wasrecovered to room temperature and stirred for another 10 hours. Thereaction solution was concentrated under reduced pressure to yieldyellow oil, which was dissolved in ethyl acetate, washed with saturatedsodium bicarbonate solution and then with saturated brine. The finalorganic phase was dried with magnesium sulfate, filtered, andconcentrated under reduced pressure. The concentrate was purified withsilica gel column chromatography (n-hexane:ethylacetate=10:1˜8:1˜4:1˜2:1˜1:1) to give Compound 2 (0.48 g, yield: 58%) asa white solid.

Step 2:

Compound 2 (0.352 g, 1.69 mmol) and 2-chloro-5-nitro-pyrimidine (0.270g, 1.69 mmol) were firstly dissolved in 12 mL acetonitrile, and thenpotassium carbonate (0.702 g, 5.08 mmol) was added to the solution. Thewhole reaction system was stirred for 3 hours at room temperature, andthen the reaction solvent was removed by rotary evaporation underreduced pressure. The concentrated substance was then dissolved in ethylacetate, washed with water and then with saturated brine. The finalorganic phase was dried with sodium sulfate, concentrated under reducedpressure and purified by silica gel column chromatography (hexane:ethylacetate=4:1˜3:1˜2:1˜1:1˜1:3) to give Product 3 (0.50 g, yield: 89%) as ayellow solid.

Step 3:

Compound 3 (0.500 g, 1.51 mmol) and palladium-carbon (0.16 g, massfraction: 5%) were added to a 25 ml two-necked flask, 10 mL methanol wasadded to the reaction system with slow stirring. After replacing the airin the whole reaction system with nitrogen, a hydrogen-filled balloonwith sufficient hydrogen was connected to the system, and then thenitrogen in the reaction system was replaced with hydrogen in theballoon (three times). The reaction system was stirred for 3 hours atroom temperature before terminating the reaction. The reaction solutionwas filtered with frit funnel to remove the palladium-carbon residue andresult in a brown filtrate. The filtrate was concentrated and purifiedwith silica gel column chromatography (n-hexane:ethylacetate=1:1˜1:2˜1:4˜1:6) to give Product 4 (0.45 g, yield: 100%) as ayellow solid.

Step 4:

3-(trifluoromethyl)benzoic acid (0.500 g, 2.63 mmol) was dispersed in 5mL thionyl chloride. The reaction system was heated to 80° C. andmaintained under stirring and refluxing for 1 hour, then cooled to roomtemperature. 10 mL toluene was added to the reaction liquid with slowstirring, and then the reaction solution was concentrated by rotaryevaporation under reduced pressure to yield light yellow oil. Theconcentrated substance was dissolved in 15 ml methylene chloride, andthen 5-amino-2-methyl-benzoic acid (0.478 g, 3.16 mmol) anddiisopropylethylamine (0.1 mL) were added to this solution. The reactionsystem was stirred overnight at room temperature to precipitate a largeamount of white solid. The reaction solution was concentrated underreduced pressure and dispersed in ethyl acetate, washed with saturatedammonium chloride solution and then with saturated brine. The finalorganic phase was dried with anhydrous sodium sulfate, concentratedunder reduced pressure, and purified by silica gel column chromatographyto give Product 5 (0.68 g, yield: 80%) as a white solid.

Step 5:

Compound 4 (0.263 g, 0.813 mmol) was dispersed in 3 mL thionyl chloride.The reaction system was heated to 80° C. and maintained under stirringand refluxing for 1 hour, then cooled to room temperature. 5 mL toluenewas added to the reaction solution with slow stirring, and the reactionsolution was concentrated under reduced pressure to yield brown oil. Theconcentrated substance was dissolved in 5 mL dichloromethane, thenCompound 5 (0.270 g, 0.894 mmol) and diisopropylethylamine amine (0.1mL) were added. The final reaction system was stirred overnight at roomtemperature, and the reaction solution was concentrated to solid underreduced pressure. The residue was dissolved in ethyl acetate, washedwith saturated sodium bicarbonate solution and then with saturatedbrine. The final organic phase was dried with anhydrous magnesiumsulfate and concentrated under reduced pressure, the concentratedsubstance was purified by silica gel column chromatography(n-hexane/ethyl acetate=2:1˜1:1˜1:2˜1:4) to give Compound 6 (0.451 g,yield: 92%) as a yellow solid.

Step 6:

Compound 6 (0.278 g, 0.458 mmol) was dispersed in 2 mL dichloromethane.2 mL trifluoroacetic acid was dropped into the reaction system slowlyunder stirring. The final reaction system was stirred for 1 hour at roomtemperature, and then was concentrated under reduced pressure to yield asolid. The residue was dissolved in ethyl acetate, washed with 10%sodium hydroxide solution and then with saturated brine. The finalorganic phase was dried with anhydrous magnesium sulfate andconcentrated under reduced pressure. The concentrated substance waspurified by silica gel column chromatography (n-hexane/ethylacetate=1:1˜1:2˜1:4) to give Product 7 (0.193 g, yield: 83%) as a whitesolid.

Synthetic Scheme II

Step 1:

Compound 7 (0.080 g, 0.16 mmol) was dispersed in a mixed solvent of THFand water (4 mL, 1:1 V/V), and then diisopropylethylamine (27 μL, 0.16mmol) was added. Acryloyl chloride (13 μL, 0.16 mmol) was dropped intothe reaction system slowly under stirring. The reaction solution wasstirred at room temperature for 2 hours, and then concentrated underreduced pressure. The residue was dissolved with ethyl acetate, washedwith 10% citric acid solution and then with saturated brine. The finalorganic phase was dried with anhydrous magnesium sulfate andconcentrated under reduced pressure. The concentrated substance waspurified by silica gel column chromatography (n-hexane/ethylacetate=1:1˜1:2) to give Product 8 (80 mg, yield: 89%) as a whitepowered solid.

Synthetic Scheme III

Step 1:

Glycine (1.00 g, 13.3 mmol) was dissolved in a mixed solvent ofpotassium hydroxide aqueous solution and 1,4-dioxane (40 mL, 1:1 V/V).(Boc)₂O (3.7 mL, 16.0 mmol) was added to the reaction solution. Thereaction system was stirred for 12 hours at room temperature, and thenthe reaction solution was concentrated under reduced pressure. Theconcentrate was dissolved in ethyl acetate, washed with 10% sodiumbisulfate solution and then with saturated brine. The final organicphase was dried with anhydrous sodium sulfate and concentrated underreduced pressure to give crude Product 9 (2.33 g, yield: 100%) as anoff-white solid.

Step 2:

Compound 7 (0.090 g, 0.177 mmol), Boc-protected glycine 9 (0.032 g,0.213 mmol) and HATU (0.101 g, 0.266 mmol) were dissolved in 3 mL DMF,diisopropylethylamine (44 μL, 0.266 mmol) was slowly added understirring. The reaction solution was stirred for 2 hours at roomtemperature, and then the solvent was removed by rotary evaporationunder reduced pressure. The residue was dissolved in ethyl acetate,washed with saturated sodium bicarbonate solution and then withsaturated brine. The final organic phase was dried with anhydrousmagnesium sulfate, filtered, concentrated under reduced pressure, andpurified by silica gel column chromatography (n-hexane:ethylacetate=1:1˜1:2˜1:4) to give Product 10 (0.106 g yield: 90%) as a whitesolid.

Step 3:

Compound 10 (0.102 g, 0.154 mmol) was dispersed in 2 mL dichloromethane.2 mL trifluoroacetic acid was dropped into the reaction system slowlyunder stirring. The final reaction system was stirred for 1 hour at roomtemperature, and then was concentrated under reduced pressure to yield asolid. The residue was dissolved with ethyl acetate, washed with 10%sodium hydroxide solution and then with saturated brine. The finalorganic phase was dried with anhydrous magnesium sulfate, concentratedunder reduced pressure, and dried in vacuum overnight to give Product 11(0.080 g, yield: 92%) as a white solid.

Step 4:

Compound 11 (0.050 g, 0.089 mmol) was dispersed in a mixed solvent ofTHF and water (2 mL, 1:1 V/V), and then diisopropylethylamine (18 μL,0.11 mmol) was added. Acryloyl chloride (14 μL, 0.18 mmol) was droppedinto the reaction system slowly under stirring. The reaction was stirredfor 2 hours at room temperature, and then was concentrated under reducedpressure. The residue was dissolved in ethyl acetate, washed withsaturated sodium bicarbonate solution and then with saturated brine. Thefinal organic phase was dried with anhydrous magnesium sulfate andconcentrated under reduced pressure. The concentrate was purified bysilica gel column chromatography (hexane:ethyl acetate=1:2˜1:4˜1:8˜100%EA) to give Product 12 (43 mg, yield: 79%) as a white solid.

Analysis of Btk In Vitro Inhibitory Activity

The Btk IC₅₀ of compounds disclosed herein was determined in anacellular kinase assay by the methods or similar methods as describedbelow.

Btk kinase activity was determined using a time-resolved fluorescenceresonance energy transfer (TR-FRET) methodology. Measurements wereperformed in a reaction volume of 50 μL using 96-well assay plates.Kinase enzyme, inhibitor, ATP (at the Km for the kinase), and 1 μMpeptide substrate (Biotin-AVLESEEELYSSARQ-NH₂) were incubated in areaction buffer composed of 20 mM Tris, 50 mM NaCl, MgCl₂ (5-25 mMdepending on the kinase), MnCl₂ (0-10 mM), 1 mM DTT, 0.1 mM EDTA, 0.01%bovine serum albumin, 0.005% Tween-20, and 10% DMSO at pH 7.4 for onehour. The reaction was quenched by the addition of 1.2 equivalents ofEDTA (relative to divalent cation) in 25 μL of 1× Lance buffer(Perkin-Elmer). Streptavidin-APC (Perkin-Elmer) and Eu-labeled p-Tyr100antibody (Perkin-Elmer) in 1× Lance buffer were added in a 25 μL volumeto give final concentrations of 100 nM and 2.5 nM, respectively, and themixture was allowed to incubate for one hour. The TR-FRET signal wasmeasured on a multimode plate reader with an excitation wavelength(λ_(Ex)) of 330 nm and detection wavelengths (λ_(Em)) of 615 and 665 nm.Activity was determined by the ratio of the fluorescence at 665 nm tothat at 615 nm. For each compound, enzyme activity was measured atvarious concentrations of compound. Negative control reactions wereperformed in the absence of inhibitor in replicates of six, and twono-enzyme controls were used to determine baseline fluorescence levels.IC50s were obtained using the program Batch K_(i) (Kuzmic et al. (2000),Anal. Biochem. 286:45-50).

According to the synthetic schemes I, II and III described above, theexample compounds 1-37 of the present invention were synthesized. Thespecific synthetic steps and characterization of the example compoundswere shown in the following table. During the analysis of Btk in vitroinhibitory activity, the IC₅₀ values of example compounds 1-37 of thepresent invention was measured. In addition, the IC₅₀ values are givenin the following table in the type of IC₅₀ value ranges, wherein “+++”represents IC₅₀<100 nM; “++” represents 100 nM<IC₅₀<1000 nM; “+”represents 1000 nM<IC₅₀<10000 nM.

TABLE 1 Synthesis of the compounds of Examples and Btk IC₅₀ values Ex-Effi- ample Structure Synthetic Scheme Structure Data cacy 1

Synthesized according to Synthetic Scheme II HRMS (ESI) m/z calculatedfor C₂₉H₂₄F₃N₆O₃ (M + H)⁺: 561.1862, found: 561.1859 +++ 2

Similar to Compound 1, but in step 1 of Synthetic Scheme II, acryloylchloride was replaced by 2-butenoyl chloride HRMS (ESI) m/z calculatedfor C₃₀H₂₆F₃N₆O₃ (M + H)⁺: 575.2018, found: 575.2015 +++ 3

Similar to Compound 1, but in step 1 of Synthetic Scheme II, acryloylchloride was replaced by vinyl sulfonyl chloride HRMS (ESI) m/zcalculated for C₂₈H₂₄F₃N₆O₄S (M + H)⁺: 597.1532, found: 597.1516 +++ 4

Synthesized according to Synthetic Scheme II HRMS (ESI) m/z calculatedfor C₃₁H₂₇F₃N₇O₄ (M + H)⁺: 618.2077, found: 618.2086 +++ 5

Synthesized according to Synthetic Scheme II, the difference lies inthat excess amount of acryloyl chloride (>26 μL, 0.32 mmol) was usedHRMS (ESI) m/z calculated for C₃₂H₂₆F₃N₆O₄ (M + H)⁺: 615.1968, found:615.2026 ++ 6

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 5-amino-2-methyl-benzoic acid was replaced by 5-amino-2- chlorobenzoic acid HRMS(ESI) m/z calculated for C₂₈H₂₁ClF₃N₆O₃ (M + H)⁺: 581.1316, found:581.1326 +++ 7

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 5-amino-2-methyl-benzoic acid was replaced by 5-amino-2- fluorobenzoic acid ¹HNMR(500 MHz, DMSO-d₆) δ 10.69 (s, 1H), 10.47 (s, 1H), 10.10 (s, 1H),9.68 (s, 1H), 8.80 (s, 2H), 8.33 (s, 1H), 8.29 (d, J = 7.8 Hz, 1H), 8.13(d, J = 6.0 Hz, 1H), 8.07 (s, 1H), 8.02- 7.98 (m, 2H), 7.81 (t, J = 7.8Hz, 1H), 7.44- 7.38 (m, 3H), 7.21 (t, J = 8.1 Hz, 1H), 6.48 (dd, J =10.2, 16.9 Hz, 1H), 6.26 (d, J = 17.0 Hz, 1H), 5.73 (d, J = 11.3 Hz,1H). HRMS (ESI) m/z +++ calculated for C₂₈H₂₁F₄N₆O₃ (M + H)+: 565.1611,found: 565.1624 8

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 5-amino-2-methyl-benzoic acid was replaced by 3-amino-benzoic acid ¹H NMR(400MHz, DMSO-d₆) δ 10.69 (s, 1H), 10.39 (s, 1H), 10.09 (s, 1H), 9.65 (s,1H), 8.83 (s, 2H), 8.31 (d, J = 8.0 Hz, 1H), 8.07 (s, 1H), 8.04 (d, J =1.3 Hz, 1H), 8.00 (d, J = 7.8 Hz, 1H), 7.79 (m, 2H), 7.57 (t, J = 7.9Hz, 1H), 7.41-7.37 (m, 2H), 7.21 (t, J = 8.1 Hz, 1H), 6.48 (dd, J =10.1, 16.9 Hz, 1H), 6.26 (dd, J = 2.1, 17.0 Hz, 1H), 5.73 (dd, J = 2.1,10.2 Hz, +++ 1H). HRMS (ESI) m/z calculated for C₂₈H₂₂F₃N₆O₃ (M + H)+:547.1705, found: 547.1782 9

Similar to Compound 1, but in step 5 of Synthestic Scheme I, Compound 5was replaced by 2- methyl-benzoic acid ¹H NMR(400 MHz, CD₃OD) δ 8.75 (s,1H), 8.13 (s, 1H), 7.50 (d, J = 7.6 Hz, 1H), 7.41-7.35 (m, 2H),7.31-7.28 (m, 3H), 7.24 (t, J = 8.0 Hz, 1H), 6.45 (dd, J = 10.0, 16.9Hz, 1H), 6.35 (dd, J = 1.6, 17.0 Hz, 1H), 5.76 +++ (dd, J = 1.6, 10.1Hz, 1H), 3.72 (t, J = 6.4 Hz, 2H), 2.47 (s, 3H), 1.88-1.85 (m, 2H). HRMS(ESI) m/z calculated for C₂₁H₂₀N₅O₂ (M + H)+: 374.1617, found: 374.163010

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 3-(trifluoromethyl) benzoic acid was replaced by propionic acid HRMS (ESI)m/z calculated for C₂₄H₂₄N₆NaO₃ (M + Na)⁺: 467.1808, found: 467.1823 ++11

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 3-(trifluoromethyl) benzoic acid was replaced by 2- naphthoic acid HRMS(ESI) m/z calculated for C₃₂H₂₇N₆O₃ (M + H)⁺: 543.2145, found: 543.2126+++ 12

Similar to Compound 1, but in step 4 of Synthetic Scheme I, 3-(trifluoromethyl) benzoic acid was replaced by 3,5- dimethoxy-benzoicacid HRMS (ESI) m/z calculated for C₃₀H₂₉N₆O₅ (M + H)⁺: 553.2199, found:553.2209 +++ 13

Similar to Compound 1, but in Step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 2,2- difluorobenzo[d][1,3]dioxole-5- carboxylic acid HRMS (ESI) m/z calculated forC₂₉H₂₃F₂N₆O₅ (M + H)⁺: 573.1698, found: 573.1712 +++ 14

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 4-(trifluoromethyl)benzoic acid HRMS (ESI) m/z calculated for C₂₉H₂₃F₃N₆NaO₃ (M + Na)⁺:583.1681, found: 583.1711 +++ 15

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-chlorobenzoic acidHRMS (ESI) m/z calculated for C₂₈H₂₄ClN₆O₃ (M + H)⁺: 527.1598, found:527.1576 +++ 16

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-fluorobenzoic acidHRMS (ESI) m/z calculated for C₂₈H₂₃FN₆NaO₃ (M + Na)⁺: 533.1713, found:533.1709 +++ 17

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 5-methylisoxazole-4-carboxylic acid HRMS (ESI) m/z calculated for C₂₆H₂₄N₇O₄ (M + H)⁺:498.1890, found: 498.1891 ++ 18

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-(dimethylamino)benzoic acid HRMS (ESI) m/z calculated for C₃₀H₂₉N₇NaO₃ (M + Na)⁺:558.2230, found: 558.2242 +++ 19

Similar to Compound 1, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by cyclopropanecar-boxylic acid HRMS (ESI) m/z calculated for C₂₅H₂₄N₆NaO₃ (M + Na)⁺:479.1808, found: 479.1820 ++ 20

Synthesized according to Synthetic Scheme I HRMS (ESI) m/z calculatedfor C₂₆H₂₂F₃N₆O₂ (M + H)⁺: 507.1756, found: 507.1737 +++ 21

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-chloro-5-(trifluoromethyl) benzoic acid HRMS (ESI) m/z calculated forC₂₆H₂₁ClF₃N₆O₂ (M + H)⁺: 541.1367, found: 541.1344 +++ 22

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by propionic acid ¹HNMR(500 MHz, DMSO-d₆) δ 10.30 (s, 1H), 9.55 (s, 1H), 9.30 (s, 1H), 8.74(s, 2H), 7.75 (s, 1H), 7.58 (d, J = 8.2 Hz, 1H), 7.22 (d, J = 8.3 Hz,1H), 7.09 (s, 1H), 6.89 (td, J = 7.8, 18.0 Hz, 2H), 6.19 (d, J = 7.6 Hz,1H), 5.13 (brs, 2H), 2.33 (s, 5H), 1.08 (t, J = 7.8, + 6.2 Hz, 3H). HRMS(ESI) m/z calculated for C₂₁H₂₃N₆O₂ (M + H)+: 391.1882, found: 391.191323

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 2-naphthoic acid ¹HNMR(400 MHz, DMSO-d₆) δ 10.56 (s, 1H), 10.40 (s, 1H), 9.34 (s, 1H), 8.79(s, 2H), 8.62 (s, 1H), 8.11-8.02 (m, 5H), 7.89 (d, J = 8.3 Hz, 1H), 7.66(s, 2H), 7.34 (d, J = 10.5 Hz, 1H), 7.11 (s, 1H), 6.94-6.86 (m, 2H),6.20 (d, J = 7.4 Hz, 1H), 5.16 (brs, 2H), 2.40 (s, 3H). HRMS (ESI) m/zcalculated for C₂₉H₂₅N₆O₂ (M + H)+: 489.2039, found: 489.2048 +++ 24

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3,5- dimethoxy-benzoicacid HRMS (ESI) m/z calculated for C₂₇H₂₇N₆O₄ (M + H)⁺: 499.2094, found:499.2096 +++ 25

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 2,2-difluoro-benzo[d][1,3]dioxole- 5-carboxylic acid HRMS (ESI) m/z calculated forC₂₆H₂₁F₂N₆O₄ (M + H)⁺: 519.1592, found: 519.1580 +++ 26

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 4-(trifluoromethyl)benzoic acid HRMS (ESI) m/z calculated for C₂₆H₂₂F₃N₆O₂ (M + H)⁺:507.1756, found: 507.1754 +++ 27

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-chlorobenzoic acid ¹HNMR(500 MHz, DMSO-d₆) δ 10.43 (s, 1H), 10.34 (s, 1H), 9.27 (s, 1H), 8.77(s, 2H), 8.04 (s, 1H), 7.94 (s, 2H), 7.82 (d, J = 8.2 Hz, 1H), 7.67 (d,J = 8.0 Hz, 1H), 7.58 (t, J = 7.8 Hz, 1H), 7.31 (d, J = 8.4 Hz, 1H),7.08 (s, 1H), 6.92-6.85 (m, 2H), 6.19 (d, J = 7.6 Hz, 1H), 4.92 (s, 2H),2.39 (s, 3H). HRMS (ESI) m/z calculated for +++ C₂₅H₂₂ClN₆O₂ (M + H)+:473.1493, found: 473.1519 28

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-fluorobenzoic acid ¹HNMR(400 MHz, DMSO-d₆) δ 10.44 (s, 1H), 10.38 (s, 1H), 9.32 (s, 1H), 8.77(s, 2H), 7.95 (d, J = 2.1 Hz, 1H), 7.85-7.78 (m, 3H), 7.64-7.58 (m, 1H),7.49-7.44 (m, 1H), 7.32 (d, J = 8.4 Hz, 1H), 7.08 (s, 1H), 6.92-6.84 (m,2H), 6.18 (d, J = 7.6 Hz, 1H), 4.96 (s, 2H), 2.39 (s, 3H). HRMS (ESI)m/z calculated for ++ C₂₅H₂₂FN₆O₂ (M + H)+: 457.1788, found: 457.1844 29

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 5-methylisoxazole-4-carboxylic acid ¹H NMR(400 MHz, DMSO-d₆) δ 10.36 (s, 1H), 10.15 (s, 1H),9.32 (s, 1H), 9.09 (s, 1H), 8.76 (s, 2H), 7.85 (d, J = 1.9 Hz, 1H), 7.73(dd, J = 2.0, 8.2 Hz, 1H), 7.31 (d, J = 8.4 Hz, 1H), 7.08 (s, 1H),6.92-6.84 (m, 2H), 6.18 (d, J = 7.6 Hz, 1H), 4.96 (s, 2H), 2.70 (s, 3H),2.38 (s, 3H). HRMS (ESI) m/z calculated for ++ C₂₅H₂₂N₇O₃ (M + H)+:444.1784, found: 444.1823 30

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-(dimethylamino)benzoic acid ¹H NMR(400 MHz, DMSO-d₆) δ 10.53 (s, 1H), 10.38 (s, 1H),9.33 (s, 1H), 8.80 (s, 2H), 8.00 (s, 1H), 7.86 (d, J = 8.3 Hz, 1H),7.34-7.27 (m, 4H), 7.09 (s, 1H), 6.94-6.84 (m, 3H), 6.18 (d, J = 7.5 Hz,1H), 4.97 (s, 2H), 2.98 (s, 6H), 2.39 (s, 3H). HRMS (ESI) m/z calculatedfor C₂₇H₂₈N₇O₂ (M + H)+: 482.2304, found: 482.2508 +++ 31

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by cyclo-propanecarboxylic acid ¹H NMR(400 MHz, DMSO-d₆) δ 10.30 (d, J = 5.4 Hz,2H), 9.32 (s, 1H), 8.75 (s, 2H), 7.78 (s, 1H), 7.58 (d, J = 8.2 Hz, 1H),7.22 (d, J = 8.4 Hz, 1H), 7.07 (s, 1H), 6.91-6.83 (m, 2H), 6.17 (d, J =7.6 Hz, 1H), 4.96 (s, 2H), 2.33 (s, 3H), 1.78 (t, J = 5.8 Hz, 1H), 0.80(t, J = 4.2 Hz, 4H). HRMS (ESI) m/z + calculated for C₂₂H₂₃N₆O₂ (M +H)+: 403.1882, found: 403.2030 32

Similar to Compound 20, but in step 4 of Synthetic Scheme I,5-amino-2-methyl- benzoic acid was replaced by 3-amino- benzoic acid ¹HNMR(400 MHz, DMSO-d₆) δ 10.70 (s, 1H), 10.36 (s, 1H), 9.32 (s, 1H), 8.78(s, 2H), 8.36-8.31 (m, 3H), 8.06 (d, J = 8.2 Hz, 1H), 7.99 (d, J = 7.8Hz, 1H), 7.84- 7.76 (m, 2H), 7.57 (t, J = 7.9 Hz, 1H), 7.09 (s, 1H),6.93-6.85 (m, 2H), 6.19 (d, J = 7.5 Hz, 1H), 4.98 (s, 2H). HRMS (ESI)m/z calculated for C₂₅H₂₀F₃N₆O₂ (M + H)+: 493.1600, found: 493.1648 ++33

Similar to Compound 20, but in step 4 of Synthetic Scheme I,5-amino-2-methyl- benzoic acid was replaced by 5-amino- 2-chlorobenzoicacid HRMS (ESI) m/z calculated for C₂₅H₁₉ClF₃N₆O₂ (M + H)⁺: 527.1210,found: 527.1193 +++ 34

Similar to Compound 20, but in step 4 of Synthetic Scheme I,5-amino-2-methyl- benzoic acid was replaced by 5-amino- 2-fluorobenzoicacid ¹H NMR(500 MHz, CD₃OD) δ 8.70 (s, 2H), 8.27 (s, 1H), 8.20 (d, J =7.9 Hz, 1H), 8.13 (dd, J = 2.7, 6.3 Hz, 1H), 7.94-7.88 (m, 2H), 7.72 (t,J = 7.8 Hz, 1H), 7.27 (t, J = 9.5 Hz, 1H), 7.20 (t, J = 2.1 Hz. 1H),7.02 (t, J = 8.0 Hz, 1H), 6.92 (dd, J = 1.0, 8.0 Hz, 1H), 6.40 (dd, J =1.3, 7.8 Hz, 1H), 4.56 (s, 1H). HRMS (ESI) m/z +++ calculated forC₂₅H₁₉F₄N₆O₂ (M + H)⁺: 511.1506, found: 511.1548 35

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by 3-(trifluoromethyl)phenyl carbamic acid HRMS (ESI) m/z calculated for C₂₆H₂₃F₃N₇O₂ (M +H)⁺: 522.1865, found: 522.1852 +++ 36

Similar to Compound 20, but in step 5 of Synthetic Scheme I, Compound 5was replaced by 2-methyl-5-(3- (trifluoromethyl) phenyl carbamoyl)benzoic acid HRMS (ESI) m/z calculated for C₂₅H₂₀F₃N₆O₂ (M + H)⁺:493.1600, found: 493.1601 ++ 37

Similar to Compound 20, but in step 4 of Synthetic Scheme I,3-(trifluoromethyl) benzoic acid was replaced by cinnamic acid(phenyl-2- acrylic acid) HRMS (ESI) m/z calculated for C₂₇H₂₅N₆O₂ (M +H)⁺: 465.2039, found: 465.2037 ++

It is understood that the examples and embodiments described herein arefor illustrative purposes only and that various modifications or changesin light thereof will be suggested to persons skilled in the art and areto be included within the spirit and purview of this application andscope of the appended claims. All publications, patents, and patentapplications cited herein are hereby incorporated by reference in theirentirety for all purposes.

What is claimed is:
 1. A compound of formula (I)

or a pharmaceutically acceptable salt thereof, wherein: W is selectedfrom H, C₁₋₆ alkyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and—(NH—CO)_(n)—NH-L-L₃; wherein: L is a bond, C₁₋₃ alkylene or C₂₋₃alkenylene; L₃ is C₃₋₈ cycloalkyl, aryl or heteroaryl, each optionallysubstituted with 1, 2 or 3 substituents selected from the groupconsisting of halogen, amino, C₁₋₆ alkyl, C₁₋₆ alkoxyl, halo-C₁₋₆ alkyl;n is an integer of 0 or 1; X is selected from H, halogen, and C₁₋₆alkyl; R₁ and R₂, same or different from each other, are eachindependently selected from H, C(O) and S(O)₂; L₁ and L₂, same ordifferent from each other, are each independently selected from C₂₋₃alkenyl optionally substituted with C₁₋₃ alkyl, and C₁₋₃alkyl-NHC(O)—C₂₋₃ alkenyl; with the provisos that when R₁ is H, L₁ isnot present; and when R₂ is H, L₂ is not present.
 2. The compound ofclaim 1, or a pharmaceutically acceptable salt thereof, wherein W isselected from H, ethyl, —(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and—(NH—CO)_(n)—NH-L-L₃, wherein: L is a bond or vinylene; L₃ iscyclopropyl, phenyl, naphthyl, isoxazolyl or benzo-[d][1,3]-dioxolegroup optionally substituted with 1 or 2 substituents selected from F,Cl, amino, methoxyl and CF₃; n is an integer of
 1. 3. The compound ofclaim 1, or a pharmaceutically acceptable salt thereof, wherein X isselected from H, F, Cl, and methyl.
 4. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein R₁ and R₂, same ordifferent from each other, are each independently selected from H, C(O)and S(O)₂; L₁ and L₂, same or different from each other, are eachindependently selected form C₂₋₃ alkenyl, and methyl-NHC(O)-ethenyl;with the provisos that when R₁ is H, L₁ is not present; and when R₂ isH, L₂ is not present.
 5. The compound of claim 1, or a pharmaceuticallyacceptable salt thereof, wherein W is selected from H, ethyl,—(NH—CO)_(n)-L-L₃, —(CO—NH)_(n)-L-L₃, and —(NH—CO)_(n)—NH-L-L₃, wherein:L is a bond or vinylene; L₃ is cyclopropyl, phenyl, naphthyl, isoxazolylor benzo-[d][1,3]-dioxole group optionally substituted with 1 or 2substituents selected from F, Cl, amino, methoxyl and CF₃; n is aninteger of 1; X is selected from H, F, Cl, and methyl; R₁ and R₂, sameor different from each other, are each independently selected from H,C(O) and S(O)₂; L₁ and L₂, same or different from each other, are eachindependently selected form C₂₋₃ alkenyl, and methyl-NHC(O)-ethenyl;with the provisos that when R₁ is H, L₁ is not present; and when R₂ isH, L₂ is not present.
 6. A compound selected from:


7. The pharmaceutical composition comprising a therapeutically effectiveamount of the compound of claim 1 and a pharmaceutically acceptableexcipient.
 8. The pharmaceutical composition comprising atherapeutically effective amount of the compound of claim 2 and apharmaceutically acceptable excipient.
 9. The pharmaceutical compositioncomprising a therapeutically effective amount of the compound of claim 3and a pharmaceutically acceptable excipient.
 10. The pharmaceuticalcomposition comprising a therapeutically effective amount of thecompound of claim 4 and a pharmaceutically acceptable excipient.
 11. Thepharmaceutical composition comprising a therapeutically effective amountof the compound of claim 5 and a pharmaceutically acceptable excipient.12. The pharmaceutical composition comprising a therapeuticallyeffective amount of the compound of claim 6 and a pharmaceuticallyacceptable excipient.
 13. The method for treating rheumatoid arthritis,comprising administering to a subject in need thereof the compound ofclaim
 1. 14. The method for treating rheumatoid arthritis, comprisingadministering to a subject in need thereof the compound of claim
 2. 15.The method for treating rheumatoid arthritis, comprising administeringto a subject in need thereof the compound of claim
 3. 16. The method fortreating rheumatoid arthritis, comprising administering to a subject inneed thereof the compound of claim
 4. 17. The method for treatingrheumatoid arthritis, comprising administering to a subject in needthereof the compound of claim
 5. 18. The method for treating rheumatoidarthritis, comprising administering to a subject in need thereof thecompound of claim
 6. 19. A method for treating rheumatoid arthritis,comprising administering to a subject in need thereof the pharmaceuticalcomposition of claim
 7. 20. The method of claim 13, wherein the subjectis a human.
 21. The method of claim 14, wherein the subject is a human.22. The method of claim 15, wherein the subject is a human.
 23. Themethod of claim 16, wherein the subject is a human.
 24. The method ofclaim 17, wherein the subject is a human.
 25. The method of claim 18,wherein the subject is a human.
 26. The method of claim 19, wherein thesubject is a human.